The goal of this translational project is to develop T cells modified to express chimeric antigen receptors (CAR T cells) as adoptive cellular therapy for patients with metastatic melanoma. We have shown in a pilot clinical trial in chronic lymphocytic leukemia that CAR T cells are capable of unprecedented on-target clinical activity. We now seek to develop new methods to safely test the antitumor CAR T cells against new target structures in patients with melanoma. This project brings together the skills of three laboratories and a team of clinical investigators to develop, implement, and test next generation CAR T cell strategies in melanoma. Our approach will use advanced engineering using mRNA vectors combined with optimized cell culture methods as our experimental platform. In preliminary work, we have (i) developed a new technology using electroporated T cells that express optimized mRNA CARs, and (ii) shown that CAR T cells targeting c-Met, a receptor tyrosine kinase markedly overexpressed in melanoma, have potent effects against c-Met+ tumor cells in vitro and in vivo. Because CAR expression in our mRNA system is transient, potentially toxic effects could be extinguished simply by discontinuing the administration of the engineered RNA CAR T cells. The ability to limit off-target exposure by discontinuing CAR administration with this new platform also creates the opportunity to rapidly test potent RNA CAR signaling domains for antitumor activity. We hypothesize that mRNA-based c-Met-specific engineered CAR T cells will allow us to optimize potency and safety in patients with metastatic melanoma and provide clinically meaningful tumor responses. Our proposal is therefore focused on testing this promising new approach with transiently expressed CARs, permitting a more rapid, cost-effective, and an intrinsically safer approach. In Aim One, we will characterize expression and anti-melanoma effects of mRNA-based c-Met scFV engineered T cells in preclinical models. In Aim Two, we will determine the clinical and immunological impact of administering autologous c-Met RNA CAR T cells in patients with metastatic melanoma in a phase I dose escalation clinical trial.